Drug Metabolism in Drug Design and Development Basic Concepts and Practice

solvents and rapidly underwent Cope elimination leading to the corresponding
hydroxylamine metabolite M-5 and acrylic acid (Scheme 11.6) (Yuan et al.,
1996).

11.7.3 Differentiation of Molecular Ions from In-Source Fragment Ions
by the Presence of Alkali Adducts

Recently, Tong and co-workers reported the identification of unstable
metabolites of Lonafarnib using ion source temperature alteration (Tong
et al., 2006). Three major drug-derived components (A, B, and C) were
characterized followingin vitroincubation of Lonafarnib with recombinant
human CYP3A4. The mass spectra for these three metabolites, obtained on an
ion trap mass spectrometer, exhibited a prominent ion atm/z635, which was
2 Th lower than that of the molecular ion for Lonafanib (m/z= 637). The
LC/MS spectra obtained on a QSTARTMmass spectrometer at relatively low
TurboSprayTMprobe temperature (250 and 350C) provided evidence that the
protonated molecular ion for metabolite A was atm/z653 with confirmatory
Na+and K+adduct ions observed atm/z675 and 691, respectively. At higher
probe temperatures, metabolite A showed no corresponding Na+ and K+
adducts form/z635 and therefore the ion atm/z635 was most likely the
in-source fragment ion formed due to facile elimination of H 2 O. Similarly,
the true molecular ion for metabolite B was determined to be atm/z667, while
the ion atm/z635 was formed in the ion source owing to the elimination of a
molecule of methanol. The true [M+H]+for metabolite C was confirmed as
m/z635 from the existence of the Na+and K+adducts atm/z657 and 673,
respectively. The detection of the alkali metal adduct ions at low ion source
temperature facilitated the identification of the true molecular ion for the
unstable metabolites. The definitive structures (Scheme 11.7) were further
characterized from accurate mass measurement, stable isotope incorporation,
and MS/MS fragmentation studies.

O N O

OH

O N O

O OH

O NOH O

OH

SC-57461

+

Cope elimination

M-4

M-5

SCHEME 11.6 Cope elimination of SC-57461-N-oxide.

352 APPLICATION OF LIQUID CHROMATOGRAPHY/MASS SPECTROMETRY